The cholinergic anti-inflammatory pathway reduces systemic TNF via acetylcholine producing memory T-cells in the spleen. These choline acetyltransferase (ChAT) expressing T-cells are also found in the intestine, where their function is unclear. We aimed to characterize these cells in mouse and human intestine and delineate their function. We made use of the ChAT-eGFP reporter mice. CD4Cre mice were crossed to ChATfl/fl mice to achieve specific deletion of ChAT in CD4+ T-cells. We observed that the majority of ChAT expressing T-cells in the human and mouse intestine have characteristics of Th17 cells, and co-express IL17A, IL22 and RORC. The generation of ChAT expressing T-cells was skewed by dendritic cells after activation of their adrenergic receptor β2. To evaluate ChAT T-cell function, we generated CD4 specific ChAT deficient mice. CD4ChAT-/- mice showed a reduced level of epithelial antimicrobial peptides (AMP) lysozyme, defA, and ang4, which was associated with an enhanced bacterial diversity and richness in the small intestinal lumen in CD4ChAT-/- mice. We conclude that ChAT expressing T-cells in the gut are stimulated by adrenergic receptor activation on dendritic cells. ChAT expressing T-cells may function to mediate the host AMP secretion, microbial growth and expansion.